EN 15788:2021
(Main)Animal feeding stuffs: Methods of sampling and analysis - Detection and enumeration of Enterococcus (E. faecium) spp. used as feed additive
Animal feeding stuffs: Methods of sampling and analysis - Detection and enumeration of Enterococcus (E. faecium) spp. used as feed additive
This document specifies general rules for the enumeration of enterococci (E. faecium) in feeding stuffs (additives, premixtures and compound feeds excluding mineral feeds) that contain enterococci as a single microorganism component or in a mixture with other microorganisms. Applying the method to premixtures and compound feeds with critical amounts of copper demands a special procedure (see A.2). The document is not applicable to mineral feeds which are defined as complementary feeding stuffs composed mainly of minerals and containing at least 40 % crude ash (Regulation (EC) 767/2009) [4].
There are different categories of feed samples:
a) Additives containing about 1010 colony forming units (CFU)/g;
b) Premixtures containing 1011 CFU/kg;
c) Compound feeds, meal or pellets which contain about 109 CFU/kg.
Futtermittel: Probenahme- und Untersuchungs-verfahren - Nachweis und Zählung von Enterococcus spp. (E. faecium) als Futtermittelzusatzstoff
Dieses Dokument legt allgemeine Regeln für die Zählung von Enterokokken (E. faecium) in Futtermitteln (Zusatzstoffe, Vormischungen und Mischfuttermittel mit Ausnahme von mineralischen Futtermitteln) fest, die Enterokokken als einzelnen mikrobiellen Bestandteil oder in einem Gemisch mit anderen Mikroorganismen enthalten. Die Anwendung des Verfahrens auf Vormischungen und Mischfuttermittel mit kritischen Kupfermengen erfordert ein besonderes Vorgehen (siehe A.2). Dieses Dokument ist nicht anwendbar auf mineralische Futtermittel, die als Ergänzungsfuttermittel definiert sind, die hauptsächlich aus Mineralstoffen zusammengesetzt sind und mindestens 40 % Rohasche enthalten (Verordnung (EG) Nr. 767/2009) [4].
Es gibt unterschiedliche Kategorien von Futtermittelproben:
a) Zusatzstoffe, die etwa 1010 koloniebildende Einheiten (KbE/g) enthalten;
b) Vormischungen, die etwa 1011 KbE/kg enthalten;
c) Mischfuttermittel, Mehle oder Pellets, die etwa 109 KbE/kg enthalten.
Aliments des animaux: Méthodes d’échantillonnage et d’analyse - Détection et dénombrement des souches de Enterococcus (E. faecium) spp. utilisées comme additifs pour l’alimentation animale
Le présent document spécifie des règles générales pour le dénombrement d’entérocoques (E. Faecium) présents dans les aliments pour animaux (additifs, prémélanges et aliments composés, à l’exception des aliments minéraux) qui contiennent des entérocoques comme seul micro-organisme constitutif ou en mélange avec d’autres micro-organismes. L’application de la méthode aux prémélanges et aux aliments composés pour animaux présentant une teneur critique en cuivre nécessite de mettre en œuvre un mode opératoire spécial (voir Annexe A.2). Le présent document ne s’applique pas aux aliments minéraux, qui se définissent comme des aliments complémentaires constitués principalement de minéraux et contenant au moins 40 % de cendre brute (Règlement (CE) 767/2009) [4].
Il existe différentes catégories d’échantillons d’aliments pour animaux :
a) les additifs contenant environ 1010 UFC/g (UFC : unités formant colonie) ;
b) les prémélanges contenant 1011 UFC/kg ;
c) les aliments composés, farines ou granulés contenant environ 109 UFC/kg.
Krma: metode vzorčenja in analize - Določanje in štetje prisotnih Enterococcus (E. faecium) spp., uporabljenih kot krmni dodatek
Ta evropski standard določa splošna pravila za štetje enterokokov v vzorcih krme (dodatki, premiksi in krma), ki vsebujejo enterokoke (E. faecium) kot eno komponento mikroorganizma ali v kombinaciji z drugimi mikroorganizmi. Uporaba metode za krmo z visoko vsebnostjo bakra (> 400 mg/kg) zahteva poseben postopek (glej dodatek A). Ta standard se ne uporablja za mineralno krmo, ki je opredeljena kot dopolnilna krma, sestavljena predvsem iz mineralov, in vsebuje najmanj 40 % surovega pepela (Direktiva Sveta 79/373/EGS).
Obstajajo različne kategorije vzorcev krme:
a) dodatki, ki vsebujejo približno 10+10 enot, ki tvorijo kolonije (CFU)/g;
b) premiksi, ki vsebujejo 10+8 CFU/g;
c) krma, moka ali peleti, ki vsebujejo približno 10+6 CFU/g ter vključujejo popolno krmo in mlečne nadomestke.
Meja zaznavanja je opredeljena v standardu EN ISO 7218.
General Information
Relations
Standards Content (Sample)
SLOVENSKI STANDARD
01-februar-2022
Nadomešča:
SIST EN 15788:2009
Krma: metode vzorčenja in analize - Določanje in štetje prisotnih Enterococcus (E.
faecium) spp., uporabljenih kot krmni dodatek
Animal feeding stuffs: Methods of sampling and analysis - Detection and enumeration of
Enterococcus (E. faecium) spp. used as feed additive
Futtermittel: Probenahme- und Untersuchungs-verfahren - Nachweis und Zählung von
Enterococcus spp. (E. faecium) als Futtermittelzusatzstoff
Aliments des animaux: Méthodes d’échantillonnage et d’analyse - Détection et
dénombrement des souches de Enterococcus (E. faecium) spp. utilisées comme additifs
pour l’alimentation animale
Ta slovenski standard je istoveten z: EN 15788:2021
ICS:
65.120 Krmila Animal feeding stuffs
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.
EN 15788
EUROPEAN STANDARD
NORME EUROPÉENNE
November 2021
EUROPÄISCHE NORM
ICS 65.120 Supersedes EN 15788:2009
English Version
Animal feeding stuffs: Methods of sampling and analysis -
Detection and enumeration of Enterococcus (E. faecium)
spp. used as feed additive
Aliments des animaux: Méthodes d'échantillonnage et Futtermittel: Probenahme- und Untersuchungs-
d'analyse - Détection et dénombrement des souches de verfahren - Nachweis und Zählung von Enterococcus
Enterococcus (E. faecium) spp. utilisées comme spp. (E. faecium) als Futtermittelzusatzstoff
additifs pour l'alimentation animale
This European Standard was approved by CEN on 2 August 2021.
CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this
European Standard the status of a national standard without any alteration. Up-to-date lists and bibliographical references
concerning such national standards may be obtained on application to the CEN-CENELEC Management Centre or to any CEN
member.
This European Standard exists in three official versions (English, French, German). A version in any other language made by
translation under the responsibility of a CEN member into its own language and notified to the CEN-CENELEC Management
Centre has the same status as the official versions.
CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia,
Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway,
Poland, Portugal, Republic of North Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and
United Kingdom.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION
EUROPÄISCHES KOMITEE FÜR NORMUNG
CEN-CENELEC Management Centre: Rue de la Science 23, B-1040 Brussels
© 2021 CEN All rights of exploitation in any form and by any means reserved Ref. No. EN 15788:2021 E
worldwide for CEN national Members.
Contents Page
European foreword . 3
Introduction . 5
1 Scope . 6
2 Normative references . 6
3 Terms and definitions . 6
4 Principle . 6
5 Diluents and culture media . 7
5.1 Diluents . 7
5.2 Culture media . 8
6 Apparatus . 10
7 Sampling . 11
8 Preparation of the test sample . 11
9 Procedure. 11
9.1 Preparation of poured agar plates for spread plate method . 11
9.2 Preparation of culture media for pour plate method . 12
9.3 Preparation of the initial suspension and decimal dilutions . 12
9.4 Inoculation and incubation of the plates . 13
9.5 Enumeration of colonies . 14
9.6 Confirmation . 14
10 Expression of results . 15
11 Precision . 15
11.1 General . 15
11.2 Interlaboratory studies . 15
11.3 Repeatability . 16
11.4 Reproducibility . 16
12 Test report . 16
Annex A (informative) Notes on the procedure . 17
A.1 General . 17
A.2 Critical copper concentration . 17
Annex B (informative) Results of the interlaboratory studies . 18
B.1 Interlaboratory study based on bile aesculin azide agar . 18
B.2 Interlaboratory study based on Slanetz and Bartley agar . 18
Bibliography . 20
European foreword
This document (EN 15788:2021) has been prepared by Technical Committee CEN/TC 327 “Animal
feeding stuffs - Methods of sampling and analysis”, the secretariat of which is held by NEN.
This European Standard shall be given the status of a national standard, either by publication of an
identical text or by endorsement, at the latest by May 2022, and conflicting national standards shall be
withdrawn at the latest by May 2022.
Attention is drawn to the possibility that some of the elements of this document may be the subject of
patent rights. CEN shall not be held responsible for identifying any or all such patent rights.
This document supersedes EN 15788:2009.
The main changes compared to the previous edition are as follows:
— Amendment of the title;
— Extension of the scope of application to all Enterococci used as feed additive;
— Updating of normative cross references; ®
— Supplement of phosphate buffered saline with Tween 80; ®
— Addition of the option to use Tween 80 supplemented phosphate buffered saline for the
preparation of the initial suspension as well as diluent for serial dilutions;
— Addition of Slanetz and Bartley agar as selective detection medium;
-1
— Replacement of the required laboratory mixer with a rotation speed of 18 000 min to
-1
22 000 min by homogenization devices, for example according to EN ISO 7218, with a maximal
-1
requested rotation speed of 10 000 min ;
— Unification of the homogenization time for the preparation of initial suspensions to five minutes for
all feed matrices;
— Addition of the option to use a spiral plater for plating;
— Preparation of initial suspensions generally conducted with tempered tPBS;
— Addition of the pour plate method as an alternative cultivation technique;
— Addition of a procedure for the investigation of feeding stuffs containing high amounts of copper in
the informative Annex A;
— Addition of validation data derived from VDLUFA ring trials of different feeding stuff matrices using
Slanetz and Bartley agar as enumeration media;
— Adjustment of the range of accepted colony numbers for counting from '≥ 30 to ≤ 350' to '≥ 10
to ≤ 200' colonies per plate.
Any feedback and questions on this document should be directed to the users’ national standards body.
A complete listing of these bodies can be found on the CEN website.
According to the CEN-CENELEC Internal Regulations, the national standards organisations of the
following countries are bound to implement this European Standard: Austria, Belgium, Bulgaria, Croatia,
Cyprus, Czech Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland, Ireland,
Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Republic of North
Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and the United
Kingdom.
Introduction
This methodology has been developed to enumerate enterococci (E. faecium) as feed additives to enable
the European Commission to control proper labelling of animal feeding products. It was compiled first
during the EU project SMT4-CT98-2235 “Methods for the official control of probiotics used as feed
additives” [1]. It was amended with a second medium from VDLUFA method 28.2.3 “Enumeration of
Enterococcus faecium” [2]. The method is based on an extensive screening of 12 pre-selected,
commercially available media for the detection and enumeration of enterococci. The specified
methodology was validated in interlaboratory studies for E. faecium ([1], [2], [3]). It can be assumed that
the method is also suitable for other Enterococcus spp.
This method is not selective for enterococci (E. faecium) used as feed additives, but can be applied to
enumerate Enterococcus spp. in additives, premixtures and compound feeds assuming that the added
enterococci (E. faecium) are present in far higher numbers than any other enterococci.
This method is not applicable for the detection of any ubiquitous or faecal contaminants of Enterococcus
spp. in food and animal feeding stuffs.
1 Scope
This document specifies general rules for the enumeration of enterococci (E. faecium) in feeding stuffs
(additives, premixtures and compound feeds excluding mineral feeds) that contain enterococci as a single
microorganism component or in a mixture with other microorganisms. Applying the method to
premixtures and compound feeds with critical amounts of copper demands a special procedure (see A.2).
The document is not applicable to mineral feeds which are defined as complementary feeding stuffs
composed mainly of minerals and containing at least 40 % crude ash (Regulation (EC) 767/2009) [4].
There are different categories of feed samples:
a) Additives containing about 10 colony forming units (CFU)/g;
b) Premixtures containing 10 CFU/kg;
c) Compound feeds, meal or pellets which contain about 10 CFU/kg.
2 Normative references
The following documents are referred to in the text in such a way that some or all of their content
constitutes requirements of this document. For dated references, only the edition cited applies. For
undated references, the latest edition of the referenced document (including any amendments) applies.
EN ISO 6498, Animal feeding stuffs - Guidelines for sample preparation (ISO 6498)
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
ISO and IEC maintain terminological databases for use in standardization at the following addresses:
— ISO Online browsing platform: available at https://www.iso.org/obp
— IEC Electropedia: available at https://www.electropedia.org/
3.1
Enterococci
Gram-positive, catalase negative cocci, which usually occurs in pairs or short chains
Note 1 to entry: This description is based on their characteristics as used for this document.
Note 2 to entry: Enterococci are classified as aerotolerant anaerobes with the ability to reduce 2,3,5-triphenyl
tetrazolium chloride to formazan and capable of hydrolyzing aesculin at 44 °C ± 0,5 °C. Enterococci form coloni
...
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